CN112586610A

CN112586610A - Extended release formulation for antibiotic replacement therapy

Info

Publication number: CN112586610A
Application number: CN202011479237.4A
Authority: CN
Inventors: 张德奎
Original assignee: Individual
Current assignee: Individual
Priority date: 2020-12-15
Filing date: 2020-12-15
Publication date: 2021-04-02

Abstract

The essential oil product of the antibiotic substitution therapy is microencapsulated, then formed into particles, and coated with fatty acid to improve the stability of the product, and has better slow-release characteristic. Considering the immature immune system of the newborn calf or piglet, the slow release characteristic is very important for applying the newborn calf or piglet. Meanwhile, the problem of commercial application of the product caused by high volatility of the cinnamaldehyde and other essential oils (such as eugenol and thymol) is solved.

Description

Extended release formulation for antibiotic replacement therapy

Technical Field

The invention belongs to the field of feed additives, and provides an extended-release formula of an essential oil formula, and a preparation method and application thereof. The product can be applied in different animal feeds, for example in the feed of newborn calves to improve the health and performance of newborn calves, and also in the improvement of the health status of newborn piglets (by adding it to the feed of sows to indirectly affect the newborn piglets).

Background

For decades, the use of antibiotics in animal food production has become a common practice. Benefits of antibiotics include infection treatment, general disease prevention and growth promotion. The widespread use of antibiotics has been considered to be a major cause of the selection of bacterial resistance. This is partly due to the fact that only low doses of antibiotics are used in very large animal populations (Marshall BM and Levy SB.2011.food animals and antibiotics: animals on human health. Clin Microbiol Rev 24: 718-33).

According to the data of the U.S. Food and Drug administration 2014, 80% of the antibiotics are consumed by the production of Food animals (FDA.2014.2011summary report on antimicrobial solids or distributed for use in Food-producing animals Rockville, Md: FDA). There is a worldwide trend towards the elimination of antibiotics for growth promotion. In the early 2020, the United states began a voluntary action aimed at eliminating antibiotics used during the growth of meat animals, but still allowed the use of antibiotics for the treatment and prevention of disease.

The plant essential oil is oily liquid extracted from plants by distillation, and is very volatile. The natural extracted plant essential oil components, such as thymol, carvacrol, cinnamaldehyde, etc., have effective antibacterial activity. Certain research has been conducted on plant essential oils in various fields such as medicine and food.

CN201710166082.0 provides a vegetable essential oil composition for dairy cows, which comprises the following components in parts by weight: 1-8 parts of eugenol, 1-15 parts of cinnamaldehyde and a mixture of carvacrol and thymol which are treated by rumen, wherein the mixture of carvacrol and thymol which are treated by rumen comprises 1-8 parts of carvacrol, 1-8 parts of thymol and 10-40 parts of a carrier for rumen bypass. The rumen protein preparation is mainly used for replacing rumen protein in rumen, inhibiting and killing harmful bacteria in intestinal tracts of dairy cows, promoting the proliferation of beneficial bacteria, preventing the diarrhea and constipation of the dairy cows, reducing the number of somatic cells, and improving the milk yield and the milk fat rate and the milk protein rate. However, the invention is mainly carried out for adult cows, and due to the strong volatility of the essential oil (the boiling point of the essential oil is extremely low), the product of the essential oil is easy to cause the essential oil to volatilize rapidly and completely without delayed release property, so that the invention is difficult to be widely applied.

CN201710166576.9 provides a plant essential oil composition, which comprises the following raw materials in parts by weight: 1-8 parts of anise oil, 1-15 parts of cinnamaldehyde and a mixture of carvacrol and thymol subjected to rumen bypass treatment, wherein the mixture of carvacrol and thymol subjected to rumen bypass treatment comprises 1-8 parts of carvacrol, 1-8 parts of thymol and 10-40 parts of a carrier for rumen bypass. The preparation method comprises coating carvacrol and thymol with rumen-bypass carrier to obtain a product, and mixing with oleum Foeniculi and cinnamaldehyde to obtain plant essential oil composition. The rumen feed additive is mainly used for replacing rumensin in rumen, inhibiting and killing harmful bacteria in intestinal tracts of ruminants, promoting proliferation of beneficial bacteria, promoting digestion and absorption of nutrient substances in small intestines of beef cattle and mutton sheep, further improving survival rate, preventing diarrhea and constipation of beef cattle and mutton sheep, improving daily gain and feed conversion rate, and preventing yellow-label meat. However, the invention is mainly carried out on beef cattle, and due to the strong volatility of the essential oil (the boiling point of the essential oil is extremely low), the product of the invention easily causes the essential oil to be rapidly and completely volatilized, has no delayed release property, and is difficult to obtain wide application.

CN201810045756.6 discloses a poultry feed additive against salmonella, wherein the additive comprises thymol, carvacrol, cinnamaldehyde, lactic acid, butyric acid, glyceryl stearate, and silicon dioxide. The feed can block vertical transmission/horizontal transmission of Salmonella; improve the fertilization rate of breeding hens and the hatching rate of breeding eggs, reduce the death rate of young chickens and reduce the number of salmonella in poultry products (poultry meat and eggs). Synergy and multiple bacteriostasis; promoting immunity, repairing, preventing and treating; the slow release process is effective in the whole digestive tract. However, the invention is mainly carried out for poultry, and the feed additive of the invention has no slow release property, is very volatile and is difficult to be applied commercially.

CN201811450869.0 discloses a water-soluble monomolecular plant essential oil, which consists of an active ingredient and a carrier, wherein the active ingredient consists of thymol and cinnamaldehyde according to the weight ratio of 1-3:1, and the carrier is water-soluble starch with the particle size of 0.005-0.02mm and the porosity of 75-85%; the weight ratio of the active ingredient to the carrier is 1-2: 1. The essential oil can enable active ingredients of the essential oil to smoothly act on the oviduct of the laying fowl, obviously improve the laying rate of the laying fowl, increase the egg weight and improve the production performance of the fowl. However, the invention is carried out for poultry, and the product belongs to a simple mixture of plant essential oil and starch, has no delayed release property, and is only in the aspect of anti-inflammation of the fallopian tube of the laying hen.

CN201911307085.7 discloses a feed additive for preventing diarrhea, which comprises the following components in parts by weight: 3-8 parts of thymol, 6-16 parts of carvacrol, 1-10 parts of cinnamaldehyde, 1-5 parts of eugenol, 1-4 parts of limonene, 1-6 parts of sweet orange oil, 1-6 parts of clove oil, 3-9 parts of cinnamon oil, 2-8 parts of butyric acid, 10-30 parts of Chinese herbal medicine extract and 40-50 parts of carrier. The feed additive is complex in composition and cannot be released with a delay.

CN201811625615.8 discloses a food calling plant essential oil product, wherein the functional components of the food calling plant essential oil product are the following components by mass percent: 3% -7% of eugenol, 2% -6% of thymol, 1% -5% of citral, 1% -5% of rutin, 1% -5% of camellia phenol and 1% -4% of other phenols; the preparation method comprises the steps of selecting raw materials, crushing and sieving, soaking in warm water, carrying out ultrasonic-assisted enzymolysis and extraction, demulsifying, extracting, drying and removing an organic solvent. The essential oil product has complex components and complex preparation process, and the obtained product does not have the property of delayed release.

CN201610958538.2 discloses a compound essential oil premix special for sows, which comprises the following components: thymol, origanum oil, eugenol, cinnamaldehyde, eucalyptus oil, fried cowherb seed, angelica sinensis extract, soybean isoflavone, mannan oligosaccharide, fructo-oligosaccharide, tea residue powder and silicon dioxide; the process comprises the following steps of putting a plurality of essential oils into a liquid mixer, uniformly mixing, putting into an impregnator, adding the tea residue powder and the silicon dioxide, infiltrating for 30 minutes, adding into a vacuum mixer, adding the rest components, sealing and mixing for 15 minutes, bagging and sealing to obtain the tea-leaf tea. The premix is mainly used for sows, has complex components and complex preparation process, and has no delayed release property.

It can be seen that although some researches on the application of plant essential oil in the aspect of feed have been carried out in the existing researches, the whole application is mainly carried out on adult pigs, cattle or poultry, and related researches on newborn calves or piglets are lacked. On the other hand, newborn animals of meat animals (Matthew A. Firth, Patricia E. Shewen, and Douglas C. Hodgins.2005 Pactive and active components of novel animal infections. animal Health Research reviews.6(2): 143) are confronted with more serious bacterial infections and possible morbidity or mortality due to their incomplete development of the immune system, and such diseases include: bacterial diarrhea, septicemia, respiratory tract infection and navel infection. Existing neonatal breeding still requires the use of antibiotics.

Therefore, there is a need to develop alternatives to antibiotics to address mortality/morbidity and improve health in newborn animals caused by bacterial infections.

Disclosure of Invention

The invention provides an essential oil product for newborn calves, piglets and other newborn animals, aiming at the problems in the prior art, and the product has the excellent effects of promoting the health of the newborn calves or the piglets and reducing the mortality/morbidity of the newborn calves or the piglets.

The essential oil product of the invention adopts microencapsulation treatment to form particles, and then uses fatty acid coating to increase the stability of the product, and has better slow release characteristic, and simultaneously, the essential oil components and the organic acid are combined to form the synergistic effect of the essential oil components and the organic acid.

The slow-release essential oil product realizes the slow release of the essential oil components on one hand and the slow release of the organic acid combined with the essential oil components in the formula on the other hand, so that the pH value of the product can be reduced at the near end of a small intestine to prevent bacteria from breeding. In contrast to products with organic acids which are taken orally directly, the latter are absorbed rapidly once they have passed through the duodenum, i.e. they do not have the prolonged release properties of the products of the patent, and do not achieve this. Considering the immature immune system of the newborn calf or piglet, the slow release property of the essential oil product of the invention is very important for applying the product to the newborn calf or piglet. At the same time, it solves the problem of commercial application of cinnamaldehyde and other essential oils (e.g. eugenol, thymol) due to their high volatility. The extended release essential oil product of the present invention can be used as an effective replacement for antibiotic therapy in young animals of a variety of animals.

First, the present invention provides a process for the preparation of an essential oil product for extended sustained release formulations, which has a decomposition degree of not more than 50% in normal digestive environment (in a common electrolyte solution of magnesium and calcium at pH 6.8 in the presence of lipase, amylase) for 4h and can be released to 8 hours, comprising a spray-dried and roll-granulated essential oil mixture embedded in a food additive and a pharmaceutically acceptable tablet coating material,

the preparation process comprises the following steps:

step 1, spray drying

Heating and dissolving cinnamaldehyde, eugenol and thymol, mixing with the aqueous solution of modified starch, removing lumps through a screen to obtain emulsified oil drops, and performing spray drying to obtain a micron particle product;

wherein the average grain diameter of the emulsified oil drops is 300-1000 nanometers, and the average grain diameter of the micron particle products is 1-100 microns;

the inlet temperature of the spray dryer is 100-225 ℃, and the outlet temperature is 50-95 ℃;

step 2, rolling granulation

Adding sorbic acid and malic acid into the micrometer particles obtained in the step 1, mixing, and rolling and granulating to obtain 18-40 mesh particles;

step 3, coating

And (3) coating the granules obtained in the step (2) in a coating chamber by using fatty acid to obtain an essential oil product.

Further preferably, the essential oil product has a degree of decomposition of not more than 80% in a common electrolyte solution of magnesium and calcium at pH 6.8 in the presence of lipase, amylase for 8 h.

Further preferably, the weight ratio of the essential oil active components of cinnamaldehyde, eugenol and thymol in the step 1 is 1-5:1-5:1-5, or 1-2:1-2:1-2, or 1:1: 1.

Further preferably, Hydroxypropylmethylcellulose (HPMC) is added to the emulsified droplets in step 1, more preferably low viscosity hydroxypropylmethylcellulose; wherein the weight ratio of the hydroxypropyl methyl cellulose to the sum of the cinnamaldehyde, the eugenol and the thymol is 1:1-5, or 1:1.5-3, or 1: 2.4.

Further preferably, maltodextrin is added to the emulsified droplets in step 1; wherein the weight ratio of the maltodextrin to the sum of the cinnamaldehyde, the eugenol and the thymol is 1:0.8-4, or 1:1-3, or 1: 1.6.

Further preferably, the modified starch in step 1 is N-Lok modified starch; wherein the weight ratio of the modified starch to the sum of the cinnamaldehyde, the eugenol and the thymol is 1:0.1-1, or 1:0.2-0.6, or 1: 0.4.

Further preferably, the average particle size of the emulsified oil droplets in the step 1 is 400-900 nm, or 600-800 nm, and the average particle size of the micron particle product is 10-60 microns, or 12-30 microns.

Further preferably, the inlet temperature of the spray dryer in step 1 is 100-.

Further preferably, sorbic acid, malic acid, hydroxypropyl methylcellulose, croscarmellose sodium, stearic acid, and magnesium stearate are added in step 2.

Further preferably, in step 2, the weight part ratio of the micron particles to sorbic acid, malic acid, hydroxypropyl methylcellulose, croscarmellose sodium, stearic acid and magnesium stearate is 30-50: 10-30: 10-30:5-10:5-10:1-3:1, and the preferred weight part ratio is 40:20:20:8.5:8.5:2: 1.

Further preferably, the weight ratio of the particles to the fatty acid in step 3 is 60-80: 40-20.

The invention also provides an essential oil product with a sustained release effect comprising a spray-dried and roller-granulated essential oil mixture embedded in a food additive and a pharmaceutically acceptable tablet coating material, wherein the essential oil mixture is at least about 40% by weight and consists of 18-40 mesh granules, and the essential oil product has a decomposition degree of not more than 50% in a normal digestive tract environment (in the presence of lipase, amylase in a common electrolyte solution of magnesium and calcium at pH 6.8) for 4 h.

Further preferred, wherein the essential oil mixture comprises about 2-8% eugenol, about 2-8% thymol and about 2-8% cinnamaldehyde embedded in a pharmaceutically acceptable excipient and/or encapsulant.

Further preferably, the weight ratio of the active components of cinnamaldehyde, eugenol and thymol in the essential oil mixture is 1-5:1-5:1-5, or 1-2:1-2:1-2, or 1:1: 1.

The invention also provides the use of an essential oil product comprising a spray-dried and roller-granulated mixture of essential oils embedded in a food additive and a pharmaceutically acceptable tablet coating material, in a prolonged sustained release formulation, which has a degree of decomposition of not more than 50% in the normal gut environment (pH 6.8 in a common electrolyte solution of magnesium and calcium in the presence of lipase, amylase) for 4 h.

Further preferably, the essential oil product has a degree of decomposition of no more than 80% in a normal gut environment (pH 6.8 in a common electrolyte solution of magnesium and calcium in the presence of lipase, amylase) for 8 h.

It is well known in the art that essential oil products are highly evaporative and volatile. Mixing them with starch and then with fatty acids, as described in the previous patent, leads to a highly unstable product. The active ingredient will evaporate within a few hours.

The method for solving the problem is to microencapsulate the essential oil by a spray drying technology to obtain a micron particle product, and in the spray drying process of the micron particle product, the added modified starch (such as N-Lok) forms a firm film around nano emulsified oil drops of essential oil active components, namely cinnamaldehyde, eugenol and thymol.

To further enhance the shell strength of the granules, the present invention may also add the polymer hydroxypropyl methylcellulose (HPMC), or use similar polymers.

In order to further protect and increase the structural stability of the active ingredient, maltodextrin may also be added as a shell material.

This process produces microparticles (about 10 microns) with a robust shell, which is very advantageous for protecting the essential oil components from evaporation and degradation, resulting in a longer shelf life of the final product.

As an example, the essential oil components of cinnamaldehyde (100-500 g), eugenol (100-500 g) and thymol (100-500 g) were added to a jar separately. The jar was capped and gently heated (about 45 ℃) to completely dissolve the thymol in the oil. 1-10 kg of N-Lok was mixed into 7 to 70 kg of deionized water using a Silverson mixer, the N-Lok solution was mixed with the essential oil components, the mixture was kept at a speed of about 9500rpm for an additional 20 minutes before reinstalling the disintegration head and used to incorporate 240g to 2400g of HPMC K100 LV into the liquid. The mixture was then poured into a 1.18mm sieve to eliminate any small lumps and then fed into a Niro spray dryer at inlet and outlet temperatures of 100-.

The mean particle size of the emulsified oil droplets was tested to be 534 nm and the spray dried essential oil particles were 10.5 microns.

The final yield was about 85%.

Next, roll granulation

In this process, the inventors tried to directly coat the microparticles with fatty acid (putting the microparticles in the coating chamber and coating with heated fatty acid, 70% by weight of the product, 30% by weight of fatty acid). Unfortunately, the microparticles are unstable in this process, causing rapid evaporation of the essential oil.

Thus, the inventors decided to granulate the microparticles into 18-40 mesh particles. Since most of the nano essential oil components are inside the particles, they remain stable to the coating step.

Granulation was carried out in a roller compactor with the following composition.

The final step is coating.

The granules were coated with fatty acid in a coating chamber, and the inventors used different percentages of coating to ensure that the final product could be released at different times.

The invention also provides a method for feeding newborn calves or piglets, which comprises the step of providing the essential oil product with the extended sustained-release formula to the newborn calves or sows.

The invention discloses a method for breeding newborn calves, which comprises the following steps: after a newborn calf is born by a cow, the calf is removed and placed in different booths, the newborn calf is fed by milk powder, the product of the invention is added into the milk powder, and the feeding addition time is 7 days.

The piglet breeding method comprises the following steps: sows are fed in a separate shed before farrowing until weaning of the piglets, and the product of the invention is added to the feed of sows 2 to 3 days before farrowing until weaning of the piglets, about 21 to 23 days after birth. The product affects the health of piglets by being secreted into the milk.

Further, the amount of the sustained release formulation to be administered to newborn calves is about 0.5 to 2 g/day, preferably about 1 g/day.

Further, the amount of the sustained-release preparation to be supplied to the sow is 5-15 g/kg of feed, and the sow consumes about 6-8 kg of feed per day.

According to the method for feeding the newborn calf or the piglet, the navy inflammation of the newborn calf or the piglet is effectively reduced and prevented, and the death rate of the newborn calf is obviously reduced within 30 days. The umbilical cord inflammation of the calf is reduced, the piglet does not have the problem of the umbilical cord inflammation, and the death rate of the piglet is obviously reduced.

The invention combines the essential oil and the organic acid into a sustained-release preparation, combines the essential oil and the organic acid, and obtains sustained-release granules by carrying out spray drying, rolling granulation and coating on the essential oil components to achieve the optimal effect and exert the effect to the maximum extent.

The clinical trial process of the invention is as follows:

1. a newborn calf.

See the examples for details.

1 gram of the granules was mixed with milk replacer. The treatment is carried out once a day.

2. Piglets.

In 11 months in 2018, a feed ingredient test was performed in the Harker Sow unit to evaluate the effect of the essential oil product on the health and viability of piglets. The test was conducted in a modern commercial swine production facility in indiana. The test animals all had a consistent commercial porcine genetic composition.

Each group of animals contained 130 adult female pigs (sows) and their piglets. The test group sows were fed on a regular diet and added with the test feed additives two days before parturition and before weaning approximately 23 days after parturition. The control group received a normal diet. Technicians caring for sows and piglets use normal production protocols.

The feed provided to the test group was produced according to industry standard commercial swine nutrition specifications. The control and test groups had the same dietary formulations and ingredients except for the test ingredients. There was no interruption of the feeding regimen/schedule. The skilled person is aware of the testing procedure but not the treatment status of the animal. Normal production indicators were recorded during the test. The production records of both groups were entered into the metafars data system and the data exported to the test coordinator for analysis.

To balance litter size, some piglets in larger litters were transferred postnatally to a smaller number of piglet cages. Mortality for each group was calculated as follows:

(original live birth quantity-live birth quantity at weaning)/original live birth quantity x 100%

Drawings

These and other features, aspects, and advantages of the present invention will become better understood with regard to the following description, appended drawings, and accompanying claims.

Figure 1, particle size distribution plot of emulsified particles of essential oil of example 1, with particle size (microns) on the abscissa and percentage on the ordinate.

Figure 2, particle size distribution of spray dried particles of example 1, with particle size (microns) on the abscissa and percentage on the ordinate.

Figure 3, dissolution profile of the essential oil product of the extended release formulation of example 2, with time on the abscissa, hours as units of measure, and percentage of dissolution on the ordinate.

Detailed Description

While the concepts of the present invention have been illustrated and described in detail in the drawings and description herein, the results in the drawings and the description are to be regarded as illustrative in nature and not as restrictive. It being understood that only exemplary embodiments have been shown and described and that all changes and modifications that come within the spirit of the invention are desired to be protected.

Unless defined otherwise, scientific and technical terms have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

The apparatus used in the present invention comprises:

rodges box dryer, Bepex rolling compressor, animal feed pellet spray Coater. Other equipment and materials are commercially available.

The inherent disease susceptibility of calves leads to frequent use of antimicrobials. Improvements in antimicrobial management require alternative therapies to improve the health and performance of calves while reducing the need for antimicrobial agents. The inventors' original design of the product was aimed at preventing small intestinal bacterial overgrowth which can lead to severe gastrointestinal symptoms and malabsorption. Thus, the present inventors have designed a sustained release mixture consisting of essential oil compounds. These compounds have less effect on normal bacterial flora but are effective against harmful bacteria. The aim is to provide a healthy small intestine environment for the animal. For this reason, the inventors applied higher doses of essential oils than those used to promote growth. To the inventors' surprise, the high dose of essential oil and slow release formulation reduced the mortality of the animals tested.

This study investigated two alternative therapies: the effects of lactoferrin (an iron binding protein found in colostrum) and essential oil products on the growth, morbidity and mortality risk of specially fed calves. On the day of arrival at the growth facility (calves aged 3 to 7 days), calves (number n ═ 80 per treatment) were randomly assigned to 1 of 3 treatments: 1) control (not supplemented), 2) lactoferrin (1g/d in milk replacer for 7d), 3) essential oil product (1g/d in milk replacer for 21 d).

Body weight of calves was measured on the day of arrival, the same measurements were taken at ages 21 and 42 days, and mortality records were obtained before 6 weeks of age. Growth was compared using analysis of repeated measures of variance (ANOVA) and differences between diarrhea (fecal score ≧ 2 with or without depression, body temperature) and disease incidence within 3 weeks were tested using the Poisson regression model (PROC GENMOD, SAS). Neither lactoferrin nor essential oil products affected the incidence of diarrhea. However, the risk of inflammation of the navel of calves receiving the essential oil product was significantly reduced compared to the control group (RR: 0.67; 95% CI: 0.46-0.97), and the mortality rate was very low at 6 weeks of age, 4, 1 and 0 deaths in the control, lactoferrin and essential oil product treated groups, respectively.

The inventors have demonstrated that higher doses and extended release of essential oil products can reduce mortality at 6 weeks of age in newborn calves treated for only 7 days.

The product of the inventor is composed of eugenol, cinnamaldehyde and thymol, and the antibacterial activity of the eugenol, the cinnamaldehyde and the thymol has a synergistic effect compared with that of a single component. The higher dosage form is selected based on slow release and concentration in the small intestine sufficient for its activity. The essential oil components are rapidly absorbed so that they do not reach the colon and interfere with the colonic flora. The product is administered daily in admixture with a milk substitute. The essential oil compounds ingested typically peak in serum within about 1 hour. The active ingredients of the product of the inventor can continuously play the role due to the slow release effect.

Example 1

The extended release composition of this example, the essential oil product having a degree of decomposition of not more than 50% in normal gut environment (pH 6.8 in a common electrolyte solution of magnesium and calcium in the presence of lipase, amylase) for 4h, comprises a spray dried and roller granulated mixture of essential oils embedded in a food additive and a pharmaceutically acceptable tablet coating material, prepared as follows:

step 1, spray drying to prepare essential oil composition

This example provides the formulation weight percentages of essential oils and other components in exemplary raw materials for spray drying. The desired weight of each component can be readily calculated in other aliquots as one skilled in the art would readily appreciate.

Heating and dissolving cinnamaldehyde, eugenol and thymol according to the raw material proportion shown in the following table, mixing with an aqueous solution of modified starch, adding hydroxypropyl methyl cellulose and maltodextrin, removing lumps through a screen mesh to obtain emulsified oil drops, and performing spray drying to obtain a micron particle product.

Step 2, dry granulation

In this example, exemplary granules of spray-dried feed were compressed to form pellets according to the table below.

Adding sorbic acid, malic acid, hydroxypropyl methylcellulose, croscarmellose sodium, stearic acid and magnesium stearate into the spray-dried micron granules obtained in the step 1 according to the proportions shown in the following table, mixing, and performing roll-pressing granulation to obtain the granules with 18-40 meshes.

And 3, coating and coating.

The particles may be coated with various coating materials to achieve the desired extended release profile.

In the process of the present inventors, they were coated with vegetable oils.

Example 2 extended Release test

Dissolution studies were conducted in the laboratory in a fixed procedure.

A beaker containing 1 liter of a solution containing magnesium chloride (3mM), calcium chloride (8mM) and sodium chloride (0.45%) was preheated to 37 ℃ in a water bath, pH 3 HCl (0.1 equiv). Then 10 grams of the product was dispersed into the solution and the solution was gently stirred. After 1 hour of addition of lipase (5 mg%), amylase (4 mg%), the pH was adjusted to 6.8 with 1N sodium hydroxide. The dispersion of the particles in the solution was observed by naked eye. The test was repeated 3 times.

The results of the testing of the essential oil product of example 1, as shown in figure 3, the Y-axis is the dissolution of the product and the X-axis is time.

Example 3 study of essential oil products to improve the health and performance of calves.

Early management of calves (calves aged 16 or 20 weeks are marketed; Terosky et al, 1997) poses special challenges to their health and welfare. Calves travel often long distances and, mixed with other animals during transport and auctions, may increase their susceptibility to disease (Taylor et al, reviewed in 2010). Diarrhea is the leading cause of morbidity and mortality in specially fed calves (Pardon et al, 2013). Strategies to reduce morbidity and early mortality sometimes include prophylactic use of antibiotics in milk or medicated milk substitutes (MR) (Berge et al, 2003), and therefore, the veal industry uses antibacterial drugs more often than other animal industries. Improved antibiotic management requires the study of alternative therapies that reduce morbidity and mortality in specially fed calves.

Iron-binding glycoprotein lactoferrin in colostrum (rybarcczyk et al, reviewed in 2017) and the natural phenylalanine-like compound cinnamaldehyde in cinnamon bark (Chapman et al, 2017) are two antibiotic substitutes investigated for livestock.

The purpose of the present study was to study the effect of essential oils on calf health, growth and mortality. The lactoferrin-treated group was used as a positive control.

The present study was conducted on a cohort of 2 specially-reared calves (120 calves per cohort) at 1 commercial farm in ohio, from 6 to 7 months 2016, and in compliance with the institutional animal care and use committee (animal use protocol: 2015a 00000131). The calves were fed a commercial milk substitute (MR) twice daily and were fed water through a metal nipple. The calves are provided with a granular starter grain of 2 to 20 weeks of age.

A random whole block design is used. Calves were randomized to one of three treatment groups, a control group (CON) (n-80 calves), which received MR supplementation. Lactoferrin treatment group (LAC) (n ═ 80 calves) were fed once daily in the evening, and 1g of lactoferrin (Tatua cooperative dairy limited, new zealand Morrinsville, new zealand) was added to the MR for 7 days. Essential oil treatment group (CIN) (n ═ 80 calves) received 1 gram of essential oil product daily, once daily, added to the MR for 21 days.

Upon arrival at the growth facility, all calves (n-240 calves) were evaluated and then twice weekly for 6 weeks. The experimenter is generally unaware of the treatment allocation groupings of calves.

Physical health assessments include rectal temperature, and scores for diarrhea, depression, respiratory disease, navel inflammation and dehydration; for detailed scoring criteria, see pempeek et al, 2017. Body weight was measured on day of arrival, 21 days and 42 days, respectively. The appetite of each calf was recorded after overnight feeding (0 ═ normal appetite; 1 ═ slow consumption; 2 ═ 1/2 for MR refusal; 3 ═ 1/2 for MR refusal). Mortality records up to 6 weeks of age were obtained from the producers.

A health outcome is clinically considered "normal" if the stool score is 0 or 1, the depression score is 0 or 1, the navel inflammation score is 0 or 1, and the skin tent score (skin tent score) is 0. Simple diarrhea is defined as a stool score of 2 or 3 with no fever or depression. Depression is defined as a depression score of 2, 3 or 4. Fever is defined as a body temperature of 39.4 ℃ or more. Complex diarrhea is considered to be diarrhea with depression and diarrhea with fever, respectively. Respiratory disease was determined by the sum of 5 clinical signs (eye discharge or head tilt or ear position, nasal discharge, evoked or spontaneous coughing, rectal temperature); each with 4 severity levels; the respiratory abnormality score is defined as a total score of 4 or higher. Inflammation of the navel was defined as 2 or 3 points. If the skin turgor is reduced (skin tents >4s), dehydration is indicated. Anorexia was defined as an appetite score of 3.

The incidence of simple, complex diarrhea, depression, respiratory disease and dehydration was not significantly different in calves receiving lactoferrin or essential oil products compared to the control group. Although there was no significant difference in the incidence of severe simple diarrhea (fecal score of 3) for each treatment group, the risk of severe simple diarrhea was lower in essential oil calves compared to control calves (RR: 0.69; 95% CI: 0.37-1.27). The risk of navel infection in essential oil calves is also lower compared to control calves. No difference in appetite scores was seen, indicating that neither lactoferrin nor the essential oil product affected the palatability of MR. By 6 weeks of age, the reported total mortality averages 2.1%; a total of 5 (5/240) calves died or were eliminated before 6 weeks of age, with 0, 1 and 4 calves in CIN, LAC and CON treatment groups, respectively. (wherein CN: control group, CIN: essential oil group, LAC: lactoferrin group, and MR: milk substitute).

The essential oil product reduces the risk of navel inflammation compared to the control group. Inflammation of the navel is a common health problem reaching calves (Wilson et al, 2000; Pempek et al, 2017). Essential oil products have been shown to inhibit the effects of proinflammatory cytokines in vitro (Chao et al, 2008), and thus, may be an alternative to antibiotics for treatment of calf navel infections.

In conclusion, the supplementation of calves in MR with lactoferrin (1g/d, continuous 7d) or with essential oil products (1g/d, continuous 21d) did not affect growth. However, the risk of inflammation of the navel of calves receiving the essential oil product was lower compared to the control group. Mortality by 6 weeks of age was low (2.1%).

TABLE 1 results of the LAC (Lactoferrin) group and CIN (essential oil product) group

Example 4 piglet feeding

Each group of animals contained 130 adult female pigs (sows) and their piglets. The test groups were fed a normal diet and the test feed additives were added two days before delivery (egg laying) to about 23 days after weaning. The control group received a normal diet. Technicians caring for sows and piglets use normal production protocols.

The feed provided to the test group was produced according to industry standard commercial swine nutrition specifications. The control and test groups had the same dietary formulations and ingredients except for the test ingredients. There was no interruption of the feeding regimen/schedule. The skilled person is aware of the results of the test, but the status of the treatment of the animal is not seen. Normal production indicators were recorded during the test. The two sets of production records were entered into the metafiles data system and the data was exported to the test coordinator for analysis.

To balance litter size, piglets are "reared" in small liter cages immediately after birth.

Mortality of piglets was calculated as follows:

(litter size of live birth-litter size at weaning)/litter size of live birth

Note that piglets were not an independent measure, and therefore the present inventors performed the analysis in the unit of sow.

The average piglet mortality for 130 sows under control conditions was 12.57% with a standard deviation of 13.39%. In contrast, the average mortality rate for piglets in 130 sows under the treatment conditions was 9.25%, with a standard deviation of 11.98%.

To compare the average mortality, the inventors used the Wilcoxon rank-sum test (W705, p 0.018). Because p values were less than a significant level of 0.05, it can be said that there was a statistically significant difference between the mean mortality rates of the two groups.

The absolute decrease in mean mortality was 3.3% (control mean mortality-treatment mean mortality).

Claims

1. A process for the preparation of an essential oil product for extended sustained release formulation for antibiotic replacement therapy, said essential oil product having a degree of decomposition of not more than 50% in a common electrolyte solution of magnesium and calcium at pH 6.8 in the presence of lipase, amylase for 4h, said essential oil product comprising a spray dried and roller granulated mixture of essential oils embedded in a food additive and a pharmaceutically acceptable tablet coating material,

the preparation process comprises the following steps:

step 1, spray drying

step 2, rolling granulation

step 3, coating

2. The preparation process of claim 1, wherein the weight ratio of the essential oil active components of cinnamaldehyde, eugenol and thymol in the step 1 is 1-5:1-5:1-5, or 1-2:1-2:1-2, or 1:1: 1.

3. The process according to any one of claims 1 to 2, wherein the modified starch in step 1 is N-Lok modified starch; wherein the weight ratio of the modified starch to the sum of the cinnamaldehyde, the eugenol and the thymol is 1:0.1-1, or 1:0.2-0.6, or 1: 0.4.

4. An essential oil product with a sustained release effect comprising a spray-dried and roller-granulated essential oil mixture embedded in a food additive and a pharmaceutically acceptable tablet coating material, wherein the essential oil mixture is at least about 40% by weight and consists of 18-40 mesh granules, the essential oil product having a degree of decomposition of not more than 50% in a common electrolyte solution of magnesium and calcium at a pH of 6.8 in the presence of lipase, amylase for 4h, the essential oil product being capable of being used as a product for antibiotic replacement therapy.

5. The essential oil product of claim 4, wherein the weight ratio of the active ingredients of cinnamaldehyde, eugenol and thymol in the essential oil mixture is 1-5:1-5:1-5, or 1-2:1-2:1-2, or 1:1: 1.

6. Use of an essential oil product according to claim 4 or claim 5 comprising a spray-dried and roller granulated mixture of essential oils embedded in a food additive and a pharmaceutically acceptable tablet coating material, in a prolonged release formulation having a degree of decomposition of not more than 50% in the presence of lipase, amylase in a common electrolyte solution of magnesium and calcium at pH 6.8 for 4 h.

7. A sustained release formulation of a plant essential oil extract in granules comprising a spray dried mixed essential oil mixture embedded in a food additive and a pharmaceutically acceptable tablet coating material, wherein the essential oil mixture consists of at least about 40% by weight of the granules, which is capable of being a product in antibiotic replacement therapy.

8. The sustained release formulation according to claim 7 wherein the essential oil mixture comprises at least three active ingredients selected from the group consisting of eugenol, thymol, cinnamaldehyde, linalool, eucalyptus oil, carvacrol and eugenol.

9. The sustained release formulation according to claim 8 wherein the essential oil mixture comprises about 8% eugenol, about 8% thymol and about 8% cinnamaldehyde embedded in pharmaceutically acceptable excipients and encapsulants.

10. Use of an essential oil product prepared by the process of manufacture according to claims 1 to 3 or an essential oil product according to claim 4 or claim 5 or a sustained release formulation according to any one of claims 7 to 9 as an animal feed additive.